La T, Collins A M, Phillips N D, Oksa A, Hampson D J
School of Veterinary and Biomedical Sciences, Murdoch University, Murdoch, WA, Australia.
Lett Appl Microbiol. 2006 Mar;42(3):284-8. doi: 10.1111/j.1472-765X.2005.01831.x.
To develop an assay to simultaneously detect Lawsonia intracellularis, Brachyspira hyodysenteriae and Brachyspira pilosicoli in pig faeces.
A multiplex-polymerase chain reaction (M-PCR) was designed to amplify a 655-base pair (bp) portion of the L. intracellularis 16S rRNA gene, a 354-bp portion of the B. hyodysenteriae NADH oxidase gene, and a 823-bp portion of the B. pilosicoli 16S rRNA gene. Specificity was assessed using 80 strains of Brachyspira spp. and 30 other enteric bacteria. Bacterial DNA was extracted from faeces using the QIAamp DNA Stool Mini Kit. The M-PCR was tested in parallel with culture and/or PCR on 192 faecal samples from eight piggeries. Faeces also were seeded with known cell concentrations of the three pathogenic species, and the limits of detection of the M-PCR tested. The M-PCR was specific, with limits of detection of 10(2)-10(3) cells of the respective species per gram of faeces.
The M-PCR is a rapid, sensitive and specific test for detecting three important enteric bacterial pathogens of pigs.
The availability of a new diagnostic M-PCR will allow rapid detection and control of three key porcine enteric pathogens.
开发一种能同时检测猪粪便中胞内劳森菌、猪痢疾短螺旋体和结肠螺旋体的检测方法。
设计了一种多重聚合酶链反应（M-PCR），用于扩增胞内劳森菌16S rRNA基因的655个碱基对（bp）片段、猪痢疾短螺旋体NADH氧化酶基因的354 bp片段以及结肠螺旋体16S rRNA基因的823 bp片段。使用80株短螺旋体属菌株和30株其他肠道细菌评估其特异性。使用QIAamp DNA粪便微量提取试剂盒从粪便中提取细菌DNA。在来自8个猪场的192份粪便样本上，将M-PCR与培养和/或PCR同时进行测试。还在粪便中接种已知细胞浓度的这三种致病菌种，并测试M-PCR的检测限。该M-PCR具有特异性，每克粪便中各菌种的检测限为10²-10³个细胞。
M-PCR是一种用于检测猪的三种重要肠道细菌病原体的快速、灵敏且特异的检测方法。
新的诊断性M-PCR的可用性将有助于快速检测和控制三种关键的猪肠道病原体。
